Clostridium Difficile Disease Research Articles

Clinical impact of a Clostridioides (Clostridium) difficile bedside infectious disease stewardship intervention.

ObjectivesTo evaluate the clinical impact of a bedside visit to patients with a positive Clostridioides difficile test on the antimicrobial stewardship of C. difficile infection (CDI) and non-C. difficile infections.MethodsAll patients ≥18 years old with positive CDI laboratory tests hospitalized between January 2017 and August 2017 received an immediate bedside intervention that consisted mainly of checking protective measures and providing recommendations on infection control and the management of CDI and other infections.ResultsA total of 214 patients were evaluated. The infectious disease (ID) physician was the first to establish protective measures in 25.2% of the cases. In 22/29 (75.9%) cases, physicians in charge accepted ID consultant recommendations to stop CDI treatment in asymptomatic patients. Unnecessary non-CDI antibiotics were discontinued in 19.1% of the cases. ID recommendations were not accepted by physicians in charge in only 12.6% of the cases.ConclusionsA bedside rapid intervention for patients with a CDI-positive faecal sample was effective in avoiding overdiagnosis and unnecessary antibiotic treatment, optimizing anti-CDI drugs, increasing compliance with infection control measures and providing educational advice.

Clostridium difficile disease in solid organ transplant recipients: a recommended treatment paradigm.

Organ transplant recipients have an increased incidence of Clostridium difficile disease and lower clinical response rates compared with the general population. Transplant specific treatment approaches are not defined. Therefore, a review of therapeutics in the transplant population is needed. A literature review on the current therapies for C. difficile was performed focusing on the evidence in transplant recipients and immunosuppressed populations. Transplant patients warrant an aggressive approach to treatment. The authors propose a suggested treatment paradigm for therapy.

Bile salt metabolism is not the only factor contributing to Clostridioides (Clostridium) difficile disease severity in the murine model of disease

ABSTRACT Susceptibility of patients to antibiotic-associated C. difficile disease is intimately associated with specific changes to gut microbiome composition. In particular, loss of microbes that modify bile salt acids (BSA) play a central role; primary bile acids stimulate spore germination whilst secondary bile acids limit C. difficile vegetative growth. To determine the relative contribution of bile salt (BS) metabolism on C. difficile disease severity, we treated mice with three combinations of antibiotics prior to infection. Mice given clindamycin alone became colonized but displayed no tissue pathology while severe disease, exemplified by weight loss and inflammatory tissue damage occurred in animals given a combination of five antibiotics and clindamycin. Animals given only the five antibiotic cocktails showed only transient colonization and no disease. C. difficile colonization was associated with a reduction in bacterial diversity, an inability to amplify bile salt hydrolase (BSH) sequences from fecal DNA and a relative increase in primary bile acids (pBA) in cecal lavages from infected mice. Further, the link between BSA modification and the microbiome was confirmed by the isolation of strains of Lactobacillus murinus that modified primary bile acids in vitro, thus preventing C. difficile germination. Interestingly, BSH activity did not correlate with disease severity which appeared linked to alternations in mucin, which may indirectly lead to increased exposure of the epithelial surface to inflammatory signals. These data confirm the role of microbial metabolic activity in protection of the gut and highlights the need for greater understanding the function of bacterial communities in disease prevention.

Trimetoprim-sulfametoxazole in ventilator-associated pneumonia: a cohort study.

To evaluate the effectiveness of trimetoprim-sulfametoxazole (TMP-SMX) for treatment of ventilator-associated pneumonia (VAP). A retrospective cohort study including patients with VAP from 2011 to 2017. Two groups were analysed: TMP-SMX group, including patients who had received TMP-SMX (as first-line and as de-escalation), and No-TMP-SMX group, including patients who had not received TMP-SMX treatment. Primary clinical outcome was mortality at 30days from starting the antibiotic treatment (T30). Secondary outcomes were mortality at end of treatment (EoT), day survival at T30, and acquisition of multidrug-resistant bacteria during hospitalization in intensive care unit. Eighty cases of VAP were included and devised into two groups: No-TMP-SMX (31/80; 39%) and TMP-SMX (49/80; 61%). Univariate analysis showed no significant differences were found when the TMP-SMX group was compared with the No-TMP-SMX group, except for frequency of male gender (p = 0.025). No significant statistical correlations between mortality at T30 and individual factors were detected by the multivariate model. No cases of either severe allergy or Clostridium difficile disease were reported in the TMP-SMX and No-TMP-SMX groups. TMP-SMX treatment was not associated with higher mortality at EoT and T30 in comparison with the No-TMP-SMX group. TMP-SMX had a good safety profile, in terms of ecology (acquisition of MDR bacteria and Clostridium difficile disease) and clinical management (no allergy events).

Vaccination against Clostridium difficile by Use of an Attenuated Salmonella enterica Serovar Typhimurium Vector (YS1646) Protects Mice from Lethal Challenge.

Clostridium difficile disease is mediated primarily by toxins A and B (TcdA and TcdB, respectively). The receptor binding domains (RBD) of TcdA and TcdB are immunogenic, and anti-RBD antibodies are protective. Since these toxins act locally, an optimal C. difficile vaccine would generate both systemic and mucosal responses. We have repurposed an attenuated Salmonella enterica serovar Typhimurium strain (YS1646) to produce such a vaccine. Plasmid-based candidates expressing either the TcdA or TcdB RBD were screened. Different vaccine routes and schedules were tested to achieve detectable serum and mucosal antibody titers in C57BL/6J mice. When given in a multimodality schedule over 1 week (intramuscularly and orally [p.o.] on day 0 and p.o. on days 2 and 4), several candidates provided 100% protection against lethal challenge. Substantial protection (82%) was achieved with combined p.o. TcdA and TcdB vaccination alone (days 0, 2, and 4). These data demonstrate the potential of the YS1646-based vaccines for C. difficile and strongly support their further development.

Correlation between tcdB gene PCR cycle threshold and severe Clostridium difficile disease

A retrospective study, including all samples tested for Clostridium difficile from 2015 to 2018, was conducted. 222 and 199 patients were respectively classified as having a mild/moderate or severe disease. A CT ≤ 26 was significantly associated with severe disease. Furthermore, low CT values were significantly associated to older patients and leukocytosis.

Evaluation of a risk score to predict future Clostridium difficile disease using UK primary care and hospital data in Clinical Practice Research Datalink

ABSTRACT We evaluated the applicability of a Clostridium difficile infection (CDI) risk index developed for patients at hospital discharge to identify persons at high-risk of CDI in a primary care population. This retrospective observational study used data from the UK Clinical Practice Research Datalink, linked with Hospital Episodes Statistics. The risk index was based on the following patient characteristics: age, previous hospitalizations, days in hospital, and prior antibiotics use. Individual risk scores were calculated by summing points assigned to pre-defined categories for each characteristic. We assessed the association of risk factors with CDI by multivariate logistic regression. The estimated CDI incidence rate was 4/10,000 and 2/10,000 person-years in 2008 and 2012, respectively. On an index with a maximal risk of 19, a cut-off for high risk of ≥7 had sensitivity, specificity and positive predictive values of 80%, 87% and 12%, respectively. A high-risk person had a ~ 35% higher risk of CDI than a low-risk person. Multivariate risk factor analysis indicated a need to reconsider the relative risk scores. The CDI risk index can be applied to the UK primary care population and help identify study populations for vaccine development studies. Reassessing the relative weights assigned to risk factors could improve the index performance in this setting.

Clostridioides difficile Infection in Chronic Kidney Disease/End-Stage Renal Disease.

Clostridioides difficile infection (CDI) is a major health-care burden and increasingly seen in patients with chronic kidney disease (CKD) and end-stage renal disease (ESRD). Increased antibiotic use, alteration in host defenses, and gastric acid suppression are some of the etiologies for increased risk of CDI in these populations. Patients with CKD/ESRD have a higher risk of initial episode, recurrence, and development of severe CDI than those without CKD or ESRD. Diagnosis and management of CDI in patients with CKD/ESRD are similar to that in the general population. The mortality, length of stay, and health-care costs are higher in patients with CDI and CKD/ESRD. Antimicrobial stewardship with reduction in antibiotic use along with infection-control measures such as contact isolation and hand hygiene with soap and water is essential in the control and prevention of CDI in patients with CKD/ESRD.

220. Taking Off With Antimicrobial Intervention Rounds (AIR): Successes of a Pilot Stewardship Service at a Tertiary-Care VA Medical Center

BackgroundProspective audit and feedback is one of the core strategies of an antimicrobial stewardship program (ASP). Here, we hypothesized that the addition of AIR to our extant ASP would enhance appropriate use of parenteral (IV) antibiotics (ABs) on a large inpatient medical service.MethodsAdult patients on medicine wards beginning in October 2017 and not followed by the infectious diseases (ID) service were included for stewardship intervention if they were on IV ABs ≥48 hours. Recommendations were classified into: (1) duration of therapy; (2) dose adjustment; (3) IV to oral conversion; (4) adverse event prevention; (5) AB avoidance; (6) anti-pseudomonal or (7) vancomycin de-escalation; (8) AB discontinuation; (9) ID consult; (10) Δ alternative AB; (11) allergy assessment; or (12) diagnostics. Early impact of the interventions was assessed after 3 months via the Standardized Antimicrobial Administration Ratio (SAAR) and compared with the 3-month, pre-AIR period. The SAAR is used to benchmark facilities’ AB use against those of similar complexity; SAAR = 1 indicates that observed = predicted use.ResultsFor 158 interventions made, the most common syndromes were pneumonia (41%), skin and soft tissue (29.4%), and urinary tract infection (17.7%). Intervention categories other than 4, 9, and 11 had acceptance rates >85% (Figure 1). The SAAR decreased from the pre- to post-AIR period in terms of agents for: broad-spectrum use in HAI (SAAR relative ratio [RR]: 0.80, 95% CI [0.73–0.88]); MRSA (SAAR RR: 0.81, 95% CI [0.73–0.91]); and all indications (SAAR RR: 0.86, 95% CI [0.82–0.90]). During the same periods, surgical wards without AIR showed no Δ in AB use.ConclusionThe majority of AB use recommendations delivered by a pharmacist–physician stewardship team were highly accepted by medical providers and led to a 15–20% decrease in overall AB use, without adverse effect during the immediate postintervention period. Potential clinical benefits, such as decreased rates of Clostridium difficile disease, will need to be measured as the AIR program advances. It is worth noting that interventions for AB allergy assessment were least accepted by providers, possibly due to time required to comply. Design of prospective audit and feedback programs may need to address this potential deficiency. Disclosures All authors: No reported disclosures.

Current Evidence in Delivery and Therapeutic Uses of Fecal Microbiota Transplantation in Human Diseases—Clostridium difficile Disease and Beyond

The use of fecal microbiota transplantation (FMT) was first described in China in the 4th century by Ge Hong when “yellow soup,” a fecal slurry, was administered for the treatment of severe food poisoning and diarrhea, a practice that continued for centuries. Bedouin groups also consumed stools of their camels as a remedy for dysentery. FMT was also applied in veterinary medicine in Europe in the 16th century. Additional therapeutic use of human excretions was described in Europe in the 18th and 19th century and in World War II, when gut bacteria were administered to German soldiers suffering from dysentery in the North African campaign. More scientifically, Eismann, in 1958, utilized fecal transplantation via enema in 4 patients for the treatment of severe pseudomembranous colitis with success. Following this report a number of isolated cases were published describing the use of FMT by different delivery routes for the treatment of a variety of illnesses.

Current Challenges in Antibiotic Stewardship in Low- and Middle-Income Countries

ASPs are key tools in the time where an increase in the rates of bacterial resistance and cases of Clostridium difficile disease is observed worldwide, forcing a more efficient use of antibiotic therapy. We review some examples of the actual state of bacterial resistance in low- and middle-income countries (LMIC), as well as some of the proposed strategies for the establishment of ASPs in resource-constrained environments. In LMIC, there are important challenges in antimicrobial prescription; although not all inhabitants have access to adequate antibiotic therapy, the consumption of these drugs is high, and is reflected in high rates of bacterial resistance. The implementation of the strategic tools within ASP is promising and good results have been achieved, with a decrease in antimicrobial consumption. The establishment of ASP is a worldwide need, and there are currently international initiatives for its development and implementation. LMIC deal with special challenges, as they need to guarantee the access to their population, but the use must be made in an appropriate manner. Currently, high rates of resistance are reported in hospital services, but in some countries, the implementation of ASP begins to yield results. Antibiotics are drugs that have a special place in the armamentarium of modern medicine. Since their introduction, they were one of the first drugs that changed the therapeutics, with the medical practice focused on symptom control to one of curative intent. In addition to the direct positive impact they have had on the morbidity and mortality associated with infectious diseases (ID), they have allowed the development of other areas, such as intensive care, neonatal care, complex surgery, and transplants, to the extent that we know them today. Unfortunately, these wonderful drugs are threatened by the emergence of bacterial resistance (BR). Although this phenomenon is characteristic of the nature of the infectious agents, exposure to antimicrobials favors and accelerates the natural selection of microorganisms better adapted to an environment in which these molecules are found. Hence, the use of antibiotics, whether for the treatment or prevention of ID in humans or animals, or with other industrial purposes in food production, is one of the activities that currently require the establishment of measures for a rational use. If we look at the use we have given to antibiotics in the previous decades, we must recognize that we have abused them, and it is in this context that antibiotic stewardship programs (ASPs) have been developed. We understand ASP as the set of measures that can be developed for the use of antibiotics in conditions in which they are really required (in a prophylactic or therapeutic way), aiming for the use of the molecules with the narrowest antibacterial spectrum, but with activity on the causative organisms of the infection, with a duration long enough to achieve their objective, but as short as possible to minimize selective pressure. The final goal of ASP is to reduce morbidity and mortality, and preserving antibiotic utility. Challenges for ASP in LMIC: 1. A high proportion of infections are caused by multidrug-resistant (MDR) microorganisms, not only in the hospital, but also in the community 2. Limitations on the diagnostic capacity for the detection of MDR microorganisms 3. Lax regulation on antibiotic use in both humans and animals 4. Deficient regulation in the food industry 5. Lack of clinician training and administration will in these issues However, proposals that try to bring together isolated efforts, such as the one set forth by the World Health Organization (WHO), have emerged in recent years. Some countries have reported their experience in the successful implementation of some of the components of ASPs, and can serve as a guide for its adaptation to local conditions. This is the basis for the protection of antibiotics, for present and future use.

Dietary trehalose enhances virulence of epidemic Clostridium difficile.

Clostridium difficile disease has recently increased to become a dominant nosocomial pathogen in North America and Europe, although little is known about what has driven this emergence. Here we show two epidemic ribotypes (RT027 and RT078) have acquired unique mechanisms to metabolize low concentrations of the disaccharide trehalose. RT027 strains contain a single point mutation in the trehalose repressor that increases this ribotype’s sensitivity to trehalose by >500 fold. Furthermore, dietary trehalose increases virulence of a RT027 strain in a mouse model of infection. RT078 strains acquired a cluster of four genes involved in trehalose metabolism, including a PTS permease that is both necessary and sufficient for growth on low concentrations of trehalose. We propose that the implementation of trehalose as a food additive into the human diet, shortly before the emergence of these two epidemic lineages, helped select for their emergence and contributed to hypervirulence.

Bovine antibodies targeting primary and recurrent Clostridium difficile disease are a potent antibiotic alternative

The increased incidence of antibiotic resistant ‘superbugs’ has amplified the use of broad spectrum antibiotics worldwide. An unintended consequence of antimicrobial treatment is disruption of the gastrointestinal microbiota, resulting in susceptibility to opportunistic pathogens, such as Clostridium difficile. Paradoxically, treatment of C. difficile infections (CDI) also involves antibiotic use, leaving patients susceptible to re-infection. This serious health threat has led to an urgent call for the development of new therapeutics to reduce or replace the use of antibiotics to treat bacterial infections. To address this need, we have developed colostrum-derived antibodies for the prevention and treatment of CDI. Pregnant cows were immunised to generate hyperimmune bovine colostrum (HBC) containing antibodies that target essential C. difficile virulence components, specifically, spores, vegetative cells and toxin B (TcdB). Mouse infection and relapse models were used to compare the capacity of HBC to prevent or treat primary CDI as well as prevent recurrence. Administration of TcdB-specific colostrum alone, or in combination with spore or vegetative cell-targeted colostrum, prevents and treats C. difficile disease in mice and reduces disease recurrence by 67%. C. difficile-specific colostrum should be re-considered as an immunotherapeutic for the prevention or treatment of primary or recurrent CDI.

Nontoxigenic Clostridium difficile: A Source for Hospital Infection?

The source of Clostridium difficile disease (CDI) in hospitals has not been resolved. In studies using whole gene sequencing (WGS), only about one third of hospital-acquired cases were genetically linked to a previous symptomatic hospital case. Confounding this issue is confusion about defining CDI. In the U.S., nucleic acid amplification tests (NAATs) alone are generally used to identify CDI in patients with diarrhea. However, increasingly, NAAT positivity is understood to indicate only carriage of the organism. …

A 10-Year Review of Total Hospital-Onset ICU Bloodstream Infections at an Academic Medical Center

The rates of central line-associated bloodstream infections (CLABSIs) in U.S. ICUs have decreased significantly, and a parallel reduction in the rates of total hospital-onset bacteremias in these units should also be expected. We report 10-year trends for total hospital-onset ICU-associated bacteremias at a tertiary-care academic medical center. This was a retrospective analysis of all positive-result blood cultures among patients admitted to seven adult ICUs for fiscal year 2005 (FY2005) through FY2014 according to Centers for Disease Control and Prevention National Healthcare Safety Network definitions. The rate of change for primary and secondary hospital-onset BSIs was determined, as was the distribution of organisms responsible for these BSIs. Data from three medical, two general surgical, one combined neurosurgical/trauma, and one cardiac/cardiac surgery adult ICU were analyzed. Across all ICUs, the rates of primary BSIs progressively fell from 2.11/1,000 patient days in FY2005 to 0.32/1,000 patient days in FY2014; an 85.0%decrease (P< .0001). Secondary BSIs also progressively decreased from 3.56/1,000 to 0.66/1,000 patient days; an 81.4%decrease (P< .0001). The decrease in BSI rates remained significant after controlling for the number of blood cultures obtained and patient acuity. An increased focus on reducing hospital-onset infections at the academic medical center since 2005, including multimodal multidisciplinary efforts to prevent central line-associated BSIs, pneumonia, Clostridium difficile disease, surgical site infections, and urinary tract infections, was associated with progressive and sustained decreases for both primary and secondary hospital-onset BSIs.

Development of a Novel Vaccine Containing Binary Toxin for the Prevention of Clostridium difficile Disease with Enhanced Efficacy against NAP1 Strains.

Clostridium difficile infections (CDI) are a leading cause of nosocomial diarrhea in the developed world. The main virulence factors of the bacterium are the large clostridial toxins (LCTs), TcdA and TcdB, which are largely responsible for the symptoms of the disease. Recent outbreaks of CDI have been associated with the emergence of hypervirulent strains, such as NAP1/BI/027, many strains of which also produce a third toxin, binary toxin (CDTa and CDTb). These hypervirulent strains have been associated with increased morbidity and higher mortality. Here we present pre-clinical data describing a novel tetravalent vaccine composed of attenuated forms of TcdA, TcdB and binary toxin components CDTa and CDTb. We demonstrate, using the Syrian golden hamster model of CDI, that the inclusion of binary toxin components CDTa and CDTb significantly improves the efficacy of the vaccine against challenge with NAP1 strains in comparison to vaccines containing only TcdA and TcdB antigens, while providing comparable efficacy against challenge with the prototypic, non-epidemic strain VPI10463. This combination vaccine elicits high neutralizing antibody titers against TcdA, TcdB and binary toxin in both hamsters and rhesus macaques. Finally we present data that binary toxin alone can act as a virulence factor in animal models. Taken together, these data strongly support the inclusion of binary toxin in a vaccine against CDI to provide enhanced protection from epidemic strains of C. difficile.

Dysbiosis in Intensive Care

The composition of the human microbiome has been recognized to have a substantial effect on multiple aspects of human health. Alterations in the microbiome are associated with diseases as diverse as Clostridium difficile disease, obesity, juvenile rheumatoid arthritis, and colon cancer, as well as an increased risk for colonization with multidrug-resistant bacteria. Noting the frequent use of antibiotic therapy and other interventions in …

Disparate prevalence of toxigenic and nontoxigenic Clostridium difficile among distinct adult patient populations in a single institution.

Clostridium difficile (CD) disease remains a costly and important hospital-associated infection. Although nontoxigenic CD is detected by some CD testing methods, can interfere with some detection algorithms and has been suggested as a treatment for CD disease, little is known about the relative occurrence of toxigenic and nontoxigenic CD in a single institution.We used both chromogenic and selective agar media to recover CD isolates and a molecular method to detect the toxin B gene from over 2400 fresh unformed stool specimens with isolates further tested for the toxin B gene. We recovered 74 nontoxigenic and 306 toxigenic CD isolates for which a collection site could be assigned.The frequency of recovery of toxigenic and nontoxigenic CD for each hospital location and the ratio of toxigenic to nontoxigenic CD were calculated. Although the overall prevalence of toxigenic and nontoxigenic CD was 12.7 % and 3.1 %, respectively, on some wards, 48 % of all CD were nontoxigenic, while on other wards, ≤5 % were nontoxigenic.The disparate ratios of nontoxigenic CD to toxigenic CD presented here for the various 'groups' within the adult veteran population are important to the ongoing discussion and reexamination of other published work on the occurrence of toxigenic and nontoxigenic CD, for evaluating the performance of CD detection tests, for designing infection control strategies and in ultimately understanding both CD carriage and disease.

Analysis of risk factors and clinical manifestations associated with Clostridium difficile disease in Serbian hospitalized patients.

Clostridium difficile is the leading cause of infectious diarrhoea in hospitalized patients. The aim of this study was to determine the risk factors important for the development of hospital-acquired Clostridium difficile-associated disease and clinical manifestations of Clostridium difficile-associated disease. The clinical trial group included 37 hospitalized patients who were selected according to the inclusion criteria. A control group of 74 hospitalized patients was individually matched with cases based on hospital, age (within 4 years), sex and month of admission.Clostridium difficile-associated disease most commonly manifested as diarrhoea (56.76%) and colitis (32%), while in 8.11% of patients, it was diagnosed as pseudomembranous colitis, and in one patient, it was diagnosed as fulminant colitis. Statistically significant associations (p < 0.05) were found with the presence of chronic renal failure, chronic obstructive pulmonary disease, cerebrovascular accident (stroke) and haemodialysis. In this study, it was confirmed that all the groups of antibiotics, except for tetracycline and trimethoprim–sulfamethoxazole, were statistically significant risk factors for Clostridium difficile-associated disease (p < 0.05). However, it was difficult to determine the individual role of antibiotics in the development of Clostridium difficile-associated disease. Univariate logistic regression also found that applying antibiotic therapy, the duration of antibiotic therapy, administration of two or more antibiotics to treat infections, administering laxatives and the total number of days spent in the hospital significantly affected the onset of Clostridium difficile-associated disease (p < 0.05), and associations were confirmed using the multivariate model for the application of antibiotic therapy (p = 0.001), duration of antibiotic treatment (p = 0.01), use of laxatives (p = 0.01) and total number of days spent in the hospital (p = 0.001). In this study of patients with hospital-acquired diarrhoea, several risk factors for the development of Clostridium difficile-associated disease were identified.

Development of SYN-004, an oral beta-lactamase treatment to protect the gut microbiome from antibiotic-mediated damage and prevent Clostridium difficile infection

The gut microbiome, composed of the microflora that inhabit the gastrointestinal tract and their genomes, make up a complex ecosystem that can be disrupted by antibiotic use. The ensuing dysbiosis is conducive to the emergence of opportunistic pathogens such as Clostridium difficile. A novel approach to protect the microbiome from antibiotic-mediated dysbiosis is the use of beta-lactamase enzymes to degrade residual antibiotics in the gastrointestinal tract before the microflora are harmed. Here we present the preclinical development and early clinical studies of the beta-lactamase enzymes, P3A, currently referred to as SYN-004, and its precursor, P1A. Both P1A and SYN-004 were designed as orally-delivered, non-systemically available therapeutics for use with intravenous beta-lactam antibiotics. SYN-004 was engineered from P1A, a beta-lactamase isolated from Bacillus licheniformis, to broaden its antibiotic degradation profile. SYN-004 efficiently hydrolyses penicillins and cephalosporins, the most widely used IV beta-lactam antibiotics. In animal studies, SYN-004 degraded ceftriaxone in the GI tract of dogs and protected the microbiome of pigs from ceftriaxone-induced changes. Phase I clinical studies demonstrated SYN-004 safety and tolerability. Phase 2 studies are in progress to assess the utility of SYN-004 for the prevention of antibiotic-associated diarrhea and Clostridium difficile disease.